Rapamycin is a clinically useful immunosuppressant that has promising anti-tumor activities. Rapamycin specifically inhibits the function of a protein kinase called the mammalian Target of Rapamycin (mTOR); however, the molecular mechanism of inhibition is poorly understood. The mammalian Target of Rapamycin belongs to the family of phosphatidylinositol-3-kinase-related kinases (PIKKs) and plays a central role in cell growth and proliferation. Accumulating evidence suggests that mTOR is a downstream target of the PI3K/Akt signaling pathway, which is frequently deregulated in many types of cancer. While mTOR regulates the translation of mRNAs critical to G1/S progression, recent evidence also implicates mTOR in the regulation of HIF-1, a transcription factor important for the hypoxic adaptation of tumor cells. We hypothesize that the aberrant activation of the PI3K/Akt/mTOR pathway plays a critical role in the growth, proliferation, and hypoxic adaptation of tumor cells. The overall goal of this revised proposal is to improve our knowledge regarding the contribution of mTOR to cancer progression, with specific emphasis on the regulation of mTOR via signaling through PI3K/Akt. This objective will be pursued by addressing the following specific aims: (1) to define the mechanism by which Ser 2448 phosphorylation regulates mTOR function and (2) to dissect the regulation of mTOR function by the Tsc1/Tsc2 tumor suppressor complex.